Skin is the largest organ of the human body. It serves as a barrier to protect against infection, toxins, microbes, and radiation. Disorders of skin not only compromise these functions, but also cause significantly psychological, social, and occupational problems. A significant portion of the world's population is afflicted with skin problems. Disorders of the skin and/or subcutaneous tissue are coded in ICD-10 (International Statistical Classification of Diseases and Related Health Problems—10th Revision), Chapter XII, and includes e.g. atopic dermatitis, psoriasis, rosacea, acne, pityriasis rosea, urticaria, erythema, and pruritus. These disorders account for a large portion of annual healthcare costs, in addition to non-financial costs, such as intractable itching, sleep deprivation, psychiatric co-morbidities time spent in treatment, inconvenience, and associated social stigma. Children with moderate to severe atopic dermatitis (AD) consistently rate their condition as having an impact on quality of life that is comparable to insulin-dependent diabetes [1]. There is a need for treatment of skin disorders. Many of these skin disorders are associated with various degree of inflammation and itch. Inflammation is a tightly regulated process aimed to eliminate intruding pathogens and remove damaged cells. The concerted action of professional phagocytes, such as macrophages, monocytes, neutrophils and certain dendritic cells, is essential to effectively clear the site of dying cells and invading pathogens as well as to restore homeostasis [2]. Dendritic cells (DC) are potent antigen presenting cells with capacity to prime naïve T cells after uptake of antigens but are also involved in keeping tolerance [3]. The functional outcome of DC action is dictated by differential expression of co-stimulatory receptors and inhibitory receptors as well as patterns of cytokine/chemokine secretion. The healthy human skin harbors at least three DC populations: Langerhans cells (LCs) in the epidermis; and interstitial CD1a+ and CD14+ DCs in the dermis [4, 5].
Oligonucleotides are short DNA or RNA molecules, oligomers, that have a wide range of applications. “CpG oligonucleotides” (or CpG-ssON) are short single-stranded synthetic DNA or RNA molecules that contain a cytosine triphosphate nucleotide (“C”) followed by a guanine triphosphate nucleotide (“G”). It is known in the art that CpG-containing nucleic acids stimulate the immune system and can be used to treat infectious diseases, allergy, asthma and other disorders. The CpG sequence in ssDNA-ODN ligands has been shown to be indispensable for activation of Toll-like receptor 9 (TLR9), which plays a fundamental role in pathogen recognition and activation of innate immunity. The stimulatory effect of the ligand is lost when the CpG repeats are removed. Consequently, the TLR-mediated immunostimulatory effect has not been shown in single-stranded oligonucleotides lacking CpG motifs (“non-CpG ssON”).
It has been shown that stimulation of the immune system with CpG-containing immunostimulatory motifs leads to induction of pro-inflammatory responses accompanied with induction of IL-10 (see examples in U.S. Pat. No. 7,807,803 B2). The anti-inflammatory cytokine IL-10 is well known for its contribution in restoration of homeostasis after cellular injury. Numerous studies in mice have shown that IL-10 is important to limit autoimmune pathologies. IL-10 has been attributed many functions including repression of the major pro-inflammatory cytokines IL-1, IL-6, IL-12 and TNF-α as well as chemokines of both the CC and CXC type [6]. The soluble ILT-6, with anti-inflammatory effects was shown to be up-regulated by IL-10 [7]. Even though the anti-inflammatory effects of IL-10 have been known for a long time and many conditions could be improved by inducing IL-10, there have been difficulties with the attempts to develop therapeutics based on the administration of IL-10. Clinical trials in humans using recombinant IL-10 have shown only marginal success [6].
It has been disclosed (Duffy et al., US2008/0299138 and WO2008/147956; Ranjith-Kumar, C. T. et al. 2008. Molecular and cellular biology 28:4507-4519) that single-stranded DNAs can be used to regulate the inflammatory response through Toll-like receptor 3 (TLR3). It has also been shown by Sköld et al. [8] that single-stranded DNA oligonucleotides (ssONs) inhibit TLR3-mediated responses in human monocyte-derived dendritic cells and in vivo in cynomolgus macaques. TLR3 is a key receptor for recognition of double-stranded RNA and initiation of immune responses against viral infections. However, hyperactive responses can have adverse effects, such as virus-induced asthma. It was shown [8] that human monocyte-derived dendritic cells up-regulate maturation markers and secrete proinflammatory cytokines on treatment with the synthetic TLR3 ligand polyinosine-polycytidylic acid (Poly(I:C)). Poly(I:C) is a synthetic agonist to for example TLR3 and is often used as an adjuvant in vaccines [see e.g. ref. 28]. It is also well known that injection of Poly(I:C) leads to an inflammatory response, for example if administered to the skin [10]. It was shown [8] that TLR3-mediated events were inhibited in cultures with CpG ssON. Poly(I:C) activation of non-hematopoietic cells was also inhibited by CpG ssON. The uptake of Poly(I:C) into cells was reduced in the presence of CpG ssON, preventing TLR3 engagement from occurring. In cynomolgus macaques, the levels of proinflammatory cytokines in nasal secretions were reduced when ssONs were administered via the intranasal route. The ssON sequences used by Sköld et al. were:
5′-GTCGTTTTGTCGTTTTGTCGTTGTTGGTGGTGGTG-3′
(CpG ssON; SEQ ID NO: 1); and
5′-GAAGTTTTGAGGTTTTGAAGTTGTTGGTGGTGGTG-3′
(non-CpG ssON; SEQ ID NO: 2).
Today's treatment of inflammatory skin disorders often includes immunosuppressive treatments such as corticosteroids and calcineurin inhibitors [9] and is often accompanied with subsequent infections of the skin due to barrier defects and nonfunctional immune defenses. In addition, prolonged treatment with corticosteroids are associated with well know toxic side effects. The pathogenesis of skin disorders, such as for example atopic dermatitis and psoriasis, were suggested to include dysregulated IL-10 production [10-13].
Many of the skin disorders or pathologies are accompanied by pruritus, a condition involving localized or general itching. A variety of causes for the condition of pruritus are known including external and endogenous causes, localized skin disorders and systemic diseases. Itch can also be produced by a variety of chemical, mechanical, thermal and electrical stimuli [14, 15].
Generally, options for effectively treating these disorders are limited. Currently available treatment modalities for these pathologies include nonspecific topical agents such as emollients and counterirritants, topical and oral drugs such as steroids, local anesthetics and antihistamines, and physical modalities such as ultraviolet phototherapy and thermal stimulation. Some of these treatments are effective in pruritic conditions of a particular etiology, while others may show general but nonspecific benefit. It is known that many corticosteroids can relieve itch and may be effective in treating some skin disorders. However, prolonged use of such corticosteroids is associated with both cutaneous and systemic toxic side effects and their widespread use is limited without medical supervision. Selenium sulfide, sulfur and salicylic acid or tar shampoo have also been employed to treat these skin conditions. In any event, remission of the pathology or pruritus is often slow and frequently incomplete.
Nonspecific topical preparations can act as moisturizing lotions or creams or as oil-based ointments that are occlusive and serve to soften dry skin as well as provide a protective layer. While such preparations may have valuable moisturizing and skin softening properties, they also possess undesirable effects in that they generally impart to the skin an uncomfortable feeling of warmth in addition to a sticky, oily, greasy or waxy feel. More importantly, these materials alone have little effect, if any, on reducing itching.
Hence, today's treatment is not sufficient and there is a need for selective anti-inflammatory compounds that can increase antibacterial defenses and ameliorate itch. There is a need for improved methods for the treatment or prophylaxis of medical conditions such as “disorders of the skin and/or subcutaneous tissue” as defined in ICD-10.